The preparation of chemically bonded high performance liquid chromatography (HPLC) stationary phases, generally made of either an inorganic or organic microspherical support, was first developed in the 1960s.1 In order to provide increased separation efficiencies and higher surface area per gram, the support is generally formed by means of an oil/water or a water/oil suspension polymerization of liquid microdroplets of monomers into spherical, porous, solid, micron-sized polymer beads or microbeads. The microbeads are either polymers or copolymers, for example, of styrene and divinylbenzene, and are in the form of a gel. The microbeads may have varying degrees of porosity, and may then be further derivatized to provide a specific surface chemistry. The HPLC gel serves as a tool for the separation of a wide variety of analytes. Packing the gels into steel and heavy walled plastic tubes, referred to as “columns”, allows the application of high pressures which are used to force the analyte and a solvent through the column at an increased rate.
“Gel permeation chromatography” (GPC), also known as “size exclusion chromatography (SEC)”, or gel-filtration chromatography (GFC), is a method of separation in which molecules are separated based on their size, molecular weight, or molecular weight distribution.
“Reverse phase” (RP) or affinity chromatography is a complementary method of separation based on molecular interactions. In RP, separation is accomplished by the differing affinity of the analyte for the column packing material and for the mobile phase or solvent which passes through the column. This method of separation comprises a whole series of techniques including reverse, normal, affinity, and ion exchange chromatography.
Thus, in contrast to RP, it is intended in SEC and GPC that the analyte should not have any enthalpic interactions with the stationary phase. Although GPC and SEC are often used interchangeably by those skilled in the art, GPC is the term generally applied to aqueous separations, while SEC is a broader term encompassing both organic and aqueous mobile phases. SEC comprises a separation of a group of analytes that “results from the distribution of the sample between the moving mobile phase and the stagnant portion of the mobile phase retained within the porous structure of the stationary phase.”1 Analytes of differing size can diffuse through the pore structure of the stationary phase to a different extent. Smaller molecules can more readily penetrate the pores of the packing material and thus they remain in the column longer, while larger molecules pass through the column in a more direct manner and elute earlier.
SEC was originally performed on crosslinked poly(dextran) and poly(saccharide) gels for biopolymers and water soluble synthetic polymers. Porous crosslinked poly(styrene) gels were introduced at the same time for the separation of organic soluble polymers. Modern SEC is accomplished using small, for example, micrometer-sized (elm-sized), rigid, porous particles usually formed on a polymeric or silica-based support.1,20 Determination of the molecular weight of a polymer sample is often accomplished by correlating the elution time with standards of known molecular weight and constructing a calibration curve.
The chemical structure of the polymeric supports used to accomplish SEC has historically been dominated by polystyrene divinylbenzene copolymers. This does not mean that these supports are not often applied to other chromatography applications including reverse phase and affinity chromatography (RP). Frechet and coworkers studied the effect of the percent divinyl benzene in the copolymer and found that the efficiencies of the packing material increase with the divinylbenzene content. They also reported that increasing the content of divinylbenzene results in an increase in the formation of micropores.3 A vast amount of similar work has been reported either in the creation or use of polystyrene divinylbenzene copolymers.4−17 To the authors' knowledge only one example of a reported synthesis of a pure divinylbenzene gel has been reported.13 Other polymers, especially methacrylates, have also been applied in the creation of column separatory media.18,20 
The surface modification of polystyrene divinylbenzene-based materials has also received considerable attention. The primary focus of this work has been in the creation of ion exchange resins for both cation and anion exchange.15,19 Leonard in a 1997 review stated that “only a few organic polymer-based packings modified with covalently bound polymer layers have been introduced recently.”20 He also stated that the main reason for this was the lack of a convenient handle from which to bond surface chemistries to poly(styrene) type gels. The main methods by which bonding chemistries have been introduced include chemical modification of the surface chemistry, co-polymerizations with functional monomers, radiochemical modification by graft polymerization, and radio-derivatization.22 Lloyd reported, without providing details of the synthetic method, the covalent bonding of a hydrophilic layer onto poly(styrene) beads, resulting in poly(hydroxyl) functionality. The alcohol functionality of this stationary phase was then used as a handle for the creation of affinity chromatography packings.15 Carbodiimide has been used as a catalyst for the reaction of different dinitrophenyl amino acids with poly(styrene-divinylbenzene) gels which were also applied as affinity packings.22 Dextran coatings have also been bonded to polystyrene gels.23 Another approach to forming functionalized supports has been the creation of new copolymers. Lewandowski and coworkers used a mixture of vinylphenol and divinylbenzene for this purpose.21 Commercial suppliers of SEC poly(styrene) divinylbenzene columns include American Polymer Standards, Shodex, Waters, Polymer Laboratories, and Jordi FLP.
Currently available column technology for SEC of polymers has reached a high level of separation efficiency. It is now possible to separate molecules that differ by as little as a single carbon atom for lower molecular weight materials. However, the long run times necessary for analysis remains a common problem with the use of current column technology. In order to reduce run times, attempts have been made to increase the flow rate. Lloyd reported the use of poly(Styrene) based columns for rapid separations in which he analyzed the column efficiency as a function of the flow rate. He found that an inverse relationship exists. Thus, the higher the flow rate used, the less efficient the separation became.15 
Another problem associated with using increased flow rates is a resulting high back pressure. One attempt to overcome this problem utilized columns of reduced dimensions and a new stationary phase. Whereas a typical SEC column is 30 cm×7.8 mm, the use of columns of 50 mm×20 mm (reduced length and increased width) was reported to result in lower back pressures, allowing for faster flow rates and shorter run times.24 Column resolution was still reported to decrease at higher flow rates, but the decrease in resolution appears to occur at a slower rate than that for prior art phases. An obvious drawback of the use of shorter columns is that the smaller amount of gel contained within a column of reduced dimensions also limits sample loadability.